Under non-standard reaction conditions, some restriction endonucleases are capable of cleaving sequences which are similar, but not identical, to their defined recognition sequence. This altered specificity has been termed “star activity”. It has been suggested that star activity is a general property of restriction endonucleases (Nasri, et al Nucleic Acids Res. 1986 14, 811). Consistent with this suggestion, many restriction endonucleases will cleave noncanonical sites under certain extreme conditions (e.g., high glycerol, high enzyme concentration, non-optimal buffer, prolonged reaction time and the presence of organic solvents or the presence of divalent cations that are not Mg2+).
Star activity can be problematic for cloning, in some amplification reactions, in gene expression assays (e.g., SAGE) and for gene assembly.
There is a constant need for restriction endonucleases that have reduced star activity.